Redefining feasibility in clinical trials: Collaborative approaches for improved site selection.

Background: This Site Feasibility Task Force convened to assess the complex and burdensome process of site feasibility in clinical trials. The objective was to create mutual understanding of challenges and provide suggestions for improving collaboration among sponsors, contract research organizations (CROs), and sites. Methods: The task force was composed of representatives from sponsors, CROs and sites (43 % Sites, 20 % Site Networks, 10 % Small/mid-size sponsors, 10 % Small/mid-size CROs, 10 % Large sponsors, 7 % Large CROs). The group collaborated to define the scope of the problem, identify challenges in the current process, and provide suggestions for improving the process. Results: The group found there is a need for better differentiation between the three main stages of feasibility, and the four sub-phases of Site Feasibility. The discussion brought to light emerging trends like early initiation of Site Feasibility and premature engagement of sites by CROs. To fully explain these challenges, the group analyzed the current practices and documented their downstream impact on clinical trial execution for all stakeholders. A list of best practices emerged naturally from this analysis. These findings are aggregated into short and actionable best practice guides. Conclusion: The task force suggests practical changes for the feasibility process and raises awareness of emerging trends and their associated risks. This awareness can begin to drive change in the site feasibility process, although industry-wide transformation will require new levels of collaboration, data standardization and automation tools. The potential benefits of evolving this process are significant and meaningful for more efficient and successful clinical trials.

Effects of Protein Unfolding on Aggregation and Gelation in Lysozyme Solutions.

In this work, we investigate the role of folding/unfolding equilibrium in protein aggregation and formation of a gel network. Near the neutral pH and at a low buffer ionic strength, the formation of the gel network around unfolding conditions prevents investigations of protein aggregation. In this study, by deploying the fact that in lysozyme solutions the time of folding/unfolding is much shorter than the characteristic time of gelation, we have prevented gelation by rapidly heating the solution up to the unfolding temperature (~80 °C) for a short time (~30 min.) followed by fast cooling to the room temperature. Dynamic light scattering measurements show that if the gelation is prevented, nanosized irreversible aggregates (about 10-15 nm radius) form over a time scale of 10 days. These small aggregates persist and aggregate further into larger aggregates over several weeks. If gelation is not prevented, the nanosized aggregates become the building blocks for the gel network and define its mesh length scale. These results support our previously published conclusion on the nature of mesoscopic aggregates commonly observed in solutions of lysozyme, namely that aggregates do not form from lysozyme monomers in their native folded state. Only with the emergence of a small fraction of unfolded proteins molecules will the aggregates start to appear and grow.

Water Proton NMR for In Situ Detection of Insulin Aggregates.

The need for quality control during the manufacturing and distribution of biopharmaceuticals is becoming increasingly necessary. At present, detecting drug degradation through the monitoring of active factor aggregation is accomplished through "invasive" techniques, such as size-exclusion chromatography (SEC), analytical ultracentrifugation (AUC), and so on. Unfortunately, these analytical methods require sampling the drug by opening the drug container that renders the remaining drug unusable regardless of the outcome of the test. Visual inspection, the current non-invasive quality control method is qualitative and can only detect visible particulates. Thus, it will miss sub-visible protein aggregates. In this paper, human insulin preparations were used to demonstrate that the transverse relaxation rate of water protons R2 ((1) H2 O) can serve as a sensitive and reliable indicator to detect and quantify both visible and sub-visible protein aggregates. R2 ((1) H2 O) is measured using a wide-bore low-field bench-top NMR instrument with permanent magnets. Such analysis could be carried out without opening the drug container, thus saving a drug for further use. The results suggest a novel, economical, non-destructive in situ analytical technique that allows for on-the-site quantification of protein aggregation in biopharmaceutical products.

Characterization and possible functions of a new filamentous bacteriophage from Vibrio cholerae O139.

The emergence and rapid rise to dominance of Vibrio cholerae O139 in India and Bangladesh in 1992 led to the consideration that choleraphage might serve as both a selective mechanism and a means for horizontal transmission of genetic information. A filamentous phage '493' from O139 strain AJ27-493 has been purified and partially characterized. The phage was inactive on classical biotype V. cholerae O1 but it was active on El Tor biotype strains isolated prior to 1994 when El Tor re-emerged in Bangladesh. More recent El Tor isolates were all resistant to the phage. The phage was also active on O139 strains. Unlike the filamentous ctx phi, the receptor for 493 is not TcpA. The phage genome was a 9.3 kb closed circular single-stranded molecule containing a 0.4 kb double-stranded stem supporting a 2 kb single-stranded loop. A 283 bp fragment was cloned and used as a probe in Southern hybridization, in parallel with total phage 493 DNA. These probes hybridized both chromosomally and extrachromosomally with most O139 strains, but not with O1 strains. Infection of hybridization-negative El Tor or O139 strains resulted in the presence of hybridizing loci (both plasmid and chromosomal), in the appearance of an 18 kDa protein, and in marked alterations in colonial morphology. Phage 493 is clearly distinct from other O139 choleraphages which have been described. Phage 493 DNA hybridized with an encapsulated non-O1 (O31) strain (NRT36S) which was isolated before O139 was recognized. NRT36S also produces a phage which can infect El Tor strains with low efficiency. Further studies may reveal whether bacteriophage play a role in the emergence and the territoriality of new choleragenic vibrios.